Mechanisms of Daxx-mediated apoptosis Daxx is a pro-apoptotic protein of unclear mechanism. The Daxx protein binds PML, the core component of nuclear structures known as PML Oncogenic Domains (PODs), and demonstrates transcriptional represser activity, as well as binding to Histone Deacetylases (HDACs) and DNA methyltransferases. Thus, Daxx- induced alterations in gene expression presumably explain its pro-apoptotic phenotype but the apoptosis- relevant genes targeted by Daxx have heretofore been undefined, and the molecular mechanism of Daxx- induced apoptosis has been controversial. Using DNA microarrays, we identified a core set of apoptosis-relevant genes consistently either up- or down-regulated by Daxx. Several of these Daxx target genes are known targets of NF-Kfi, including cFLIP and cIAP2, which encode caspase-inhibitory proteins. Based on these and other related observations presented in the application, we propose the hypothesis that Daxx promotes apoptosis by binding to Rel-B and silencing Rel-B target genes. We will address five specific questions as we test this hypothesis and gain a better understanding of the mechanisms by which Daxx controls apoptosis, including: (1) Determining how Daxx interacts with Rel-B and ascertaining whether this interaction is critical for the pro-apoptotic phenotype of Daxx; (2) Studying how Daxx represses Rel-B function; (3) Evaluating how the trafficking of Daxx into PODs affects its ability to associate with and suppress Rel-B; (4) Examining which Daxx target genes are important for its pro-apoptotic phenotype; and (5) Exploring how Daxx modifies cell death outcomes in the context of cell stimulation by TNF family cytokines.